Luckau Protocols

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(New page: ==Tara Luckau Protocols== Clark Laboratory ===Agarose Gel===)
(Tara Luckau Protocols)
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===Agarose Gel===
===Agarose Gel===
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====Purpose====
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Agarose gels are used to verify the presence and estimate the size of DNA (both genomic and amplified). When loaded into a gel and subjected to an electric current, DNA fragments migrate to the positive terminal. Small DNA fragments migrate more rapidly than large ones. By running a commercially available ladder of fragments of known size, we can estimate the DNA fragment size in our sample of interest.
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To visualize the DNA after electrophoresis, a dye called Gel Red is added during the agarose-making process. The dye binds to the DNA and flouresces when exposed to UV light.
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====Materials To Be Familiar With====
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* Gel Rig, caster tray, combs
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* [[Image:OwlA2Large.jpg]]  [[Image:OwlA2Large_draw.jpg|300 px]]
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====Protocol====
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# Make gel using 1x TAE and agarose
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#* Use 1% for genomic DNA, 2% for amplified DNA
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{| border="1" style="margin: 1em auto 1em auto" "text-align: center"
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|-
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! Gel Rig Size !! 1x TAE (mL) !! 1.5% agarose (g) !! 2% agarose (g) !! Gel Red (µL) !! Sample volume (µL) !! max voltage (V)
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|-
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| small || 50 || _ || 1 || 5 || 4-6 || 80
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|-
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| rowspan="2"| medium || rowspan="2"| 130 || rowspan="2"| _ || rowspan="2"| 2.6 || rowspan="2"| 13 || 24-tooth comb: 6-10 || rowspan="2"| 120
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|-
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| 36-tooth comb: 3-5
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|-
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| large || _ || _ || _ || _ || _ || 170
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|}
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#

Revision as of 19:12, 1 November 2010

Contents

Tara Luckau Protocols

Clark Laboratory

Agarose Gel

Purpose

Agarose gels are used to verify the presence and estimate the size of DNA (both genomic and amplified). When loaded into a gel and subjected to an electric current, DNA fragments migrate to the positive terminal. Small DNA fragments migrate more rapidly than large ones. By running a commercially available ladder of fragments of known size, we can estimate the DNA fragment size in our sample of interest.

To visualize the DNA after electrophoresis, a dye called Gel Red is added during the agarose-making process. The dye binds to the DNA and flouresces when exposed to UV light.

Materials To Be Familiar With

  • Gel Rig, caster tray, combs
  • Image:OwlA2Large.jpg

Protocol

  1. Make gel using 1x TAE and agarose
    • Use 1% for genomic DNA, 2% for amplified DNA
Gel Rig Size 1x TAE (mL) 1.5% agarose (g) 2% agarose (g) Gel Red (µL) Sample volume (µL) max voltage (V)
small 50 _ 1 5 4-6 80
medium 130 _ 2.6 13 24-tooth comb: 6-10 120
36-tooth comb: 3-5
large _ _ _ _ _ 170
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